The ex vivo cultivation of aspirated organ-function cells or of cells with a partial organ function, like human tissue, bone marrow, liver cells and skin cells, is of great interest in the medicinal, therapeutical and pharmaceutical and research and development fields. For example, various clinical treatments utilize bone marrow, umbilical cord blood or blood recovered from peripheral stem cells.
Of special interest is the field of leukemia treatment for which the cultivation of bone marrow stem cells is important for increasing or at least maintaining levels of such cells in the treated organism or maintaining equilibria involving same so as to overcome the donor problem, problems of acceptance of foreign cells, rejection, etc. In the past, efforts have been made to overcome these problems in various ways.
For example in an autologous bone marrow transplant techniques, bone marrow is removed from the patient/recipient and returned to the patient after, for instance, cancer therapy.
In the future, therapy is considered in which the leukemic or tumor infiltrated bone marrow is expected to be subjected externally of the patient to a treatment or healing and then returned to the patient (ex vivo purging).
In allogenic bone marrow transplant techniques, the healthy bone marrow of a donor is administered to the leukemic patient from whom bone marrow has been removed. Complications can arise here in that the foreign bone marrow can be rejected or can treat the tissue of the patient/recipient as foreign tissue and attack it (graft vs. host reaction).
In the past, in practically all cases bone marrow had been transplanted only as a cell mixture. More recently, however, the stem cells are recovered from the bone marrow by the administration of cytokines in a process in which the cytokines mobilize the stem cells in the peripheral blood. These stem cells are purified immunologically and then transplanted to the patient. The need for the removal of bone marrow from a donor is thus eliminated along with the pain and stress on the body with which the bone marrow extraction has been associated. In the stem cell recovery procedure, only the desired stem cells are removed and the remaining cells are returned to the donor so that the loss of hemapoietic cells is a minimum for the donor.
A process for increasing or at least maintaining the concentration of such organ-function cells is important for all of the more recent treatment techniques as described above. The maintenance or cell-increasing cultivation of progenitor cells, (bone marrow stem cells) is expressly important for the production of blood cells for blood transfusions, to replace donor blood, from autologous cells from a patient himself or herself or for donor stem cells for allogenic transfusions and wherever mature blood cells are required. The latter can be utilized in transfusions requiring erythrocytes, thrombocytes and granulocytes. This can greatly reduce the danger of infection for the blood recipient greatly or eliminate it entirely and can insure a uniform product quality.
Furthermore, with cultivation of organ-function cells yielding an increase in the cells, a donor bank can be established in which all of the required specific stem cells for a transplant can be held in readiness. This can reduce the enormous expense and time pressure which can arise at the time that an organ-cell transplant is required.
For over 10 years efforts have been made to cultivate organ-function cells and mixed population cells derived from tissue. However, true long term cultures have been found to be extraordinarily problematical.
From EP 0 241 578 A1, for example, it has been proposed to cultivate bone marrow preparations, especially hemapoietic stem cells, on stroma or fiberblasts which initially have been cultured on a network especially of nylon before being inoculated with the hemapoietic cells.
According to EP 0 358 506 A1, carriers of optional configuration can be suspended in a growth medium and the culture can be carried out with periodic medium replacement.
In WO 92/21402, the ex vivo cultivation of hemapoietic precursor cells is carried out with a prior treatment or in the presence of growth factors or cytokines.
From WO 93/18136, the cultivation of hemapoietic cells on microcarrier beads is taught with continuous or intermittent growth medium supply, the process being carried out in a stirred reaction vessel or airlift bioreactor. Described here is the cultivation of cells which are grown on collagen in spinner flasks.
Finally, T.-Y. WANG et al u.a. (Ann. NY. Acad Sci. 1990, Pages 274-284) describe a three-chamber apparatus for the culturing of hemapoietic cells which are grown on highly porous collagen microspheres. The latter are provided as a packing in the intermediate chamber and the packing is separated from the chambers traversed by the growth medium by a perfusion membrane.
Even these techniques do not result in a true long term cultivation of organ-function cells.